Mutations in the VHL tumor suppressor gene cause the familial cancer syndrome, von Hippel- Lindau (VHL) disease, which is characterized by a predisposition to renal cell carcinoma, pheochromocytoma, and blood vessel tumors (hemangioblastomas). A genotype-phenotype correlation exists whereby specific VHL mutations have a higher risk of a particular subset of these tumor types. The most recognized biochemical function of VHL gene products (hereafter called pVHL) is as an integral member of an E3 ubiquitin ligase complex that targets specific substrates for ubiquitination and subsequent degradation. The degradation of one substrate, hypoxia-inducible factor alpha (HIFa) subunits, has emerged as a key pVHL function necessary for tumor suppression in renal cells. Contrary to the notion that HIFa dysregulation is solely responsible for all of the tumor types seen in VHL disease, a class of VHL mutants that are associated with a high risk of pheochromocytoma have apparently normal HIFa ubiquitination function, suggesting that these mutations affect an alternative VHL activity. In previous work, our laboratory has identified or characterized cellular roles for pVHL that are either fully or partly independent of HIFa regulation. These pVHL roles include regulation of cell differentiation, integrin expression, fibronectin deposition, and assembly of tight junctions. This project will further explore and examine these and other pVHL functions in renal and pheochromocytoma cells in order to clarify their relationship to both HIFa regulation and VHL tumor suppression. The specific aims of this proposal are: 1) To identify and examine HIFa independent VHL- associated roles which contribute to the phenotype-genotype correlation VHL disease;2) To investigate the relationship between PKC and VHL-mediated cellular phenotypes in adrenal and renal cells;and 3) To test the contribution of relative levels of HIF-1 and HIF-2 on apoptosis and apoptotic pathways in renal cell carcinoma. The proposed investigations will utilize established cell culture systems, with genetic manipulation of HIFa and pVHL levels, along with expression of mutant VHL proteins to analyze various VHL-mediated cellular phenotypes. The influence of another identified pVHL target, atypical protein kinase C, on these cellular phenotypes will be investigated similarly. The overall goal is to determine the underlying cellular mechanisms responsible for observed VHL-associated phenotypic changes and for tumorigenesis in VHL disease. It is anticipated that these studies will also provide insight into cellular differentiation and growth arrest mechanisms applicable to a wider range of tumor types and foster a better understanding of tumorigenic processes at the cellular level. PUBLIC HEALTH RELEVANCE: Disruptions of the VHL tumor suppression gene lead to kidney, adrenal, and blood vessel tumors. The research proposed in this project aims at understanding the changes that occur to cells as a result of VHL disruption, and how these changes may result in tumor formation (for these different tumor types). This information may lead to the design of better treatments and/or therapeutics for these tumors. The project should also advance our knowledge of cellular biology, providing a better understanding of some key events and processes that occur in cells, such as differentiation and growth arrest mechanisms, which may be applicable to a broader range of cancer types.